the phosphate requirement of the ferns byarizona.openrepository.com/arizona/bitstream/10150/... ·...
TRANSCRIPT
![Page 1: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/1.jpg)
The phosphate requirement of the fernsNotholaena cochisensis and Notholaena sinuata
Item Type text; Thesis-Reproduction (electronic)
Authors Ferrell, Cleon Duane, 1941-
Publisher The University of Arizona.
Rights Copyright © is held by the author. Digital access to this materialis made possible by the University Libraries, University of Arizona.Further transmission, reproduction or presentation (such aspublic display or performance) of protected items is prohibitedexcept with permission of the author.
Download date 13/05/2018 01:07:30
Link to Item http://hdl.handle.net/10150/551800
![Page 2: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/2.jpg)
THE PHOSPHATE REQUIREMENT OF THE FERNS
NOTHOLAENA COCHISENSIS AND NOTHOLAENA SINUATA
by
Cleon Duene Ferrell
A T h e s is S u b m itte d t o th e
COMMITTEE ON AGRICULTURAL BIOCHEMISTRY AND NUTRITION
I n P a r t i a l F u l f i l l m e n t o f th e R e q u ire m e n ts
F o r th e D eg ree o f
MASTER OF SCIENCE
I n th e G ra d u a te C o l le g e
THE UNIVERSITY OF ARIZONA
1965
V
![Page 3: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/3.jpg)
STATEMENT BY AUTHOR
T h is t h e s i s h a s b e e n s u b m it te d i n p a r t i a l f u l f i l l m e n t o f r e q u i r e m e n ts f o r an ad v an ced d e g re e a t The U n i v e r s i t y o f A r iz o n a and i s d e p o s i t e d i n th e U n i v e r s i t y L ib r a r y t o b e made a v a i l a b l e t o b o r ro w e rs u n d e r r u l e s o f t h e L ib r a r y .
B r i e f q u o ta t io n s from t h i s t h e s i s a r e a l lo w a b le w i th o u t s p e c i a l p e r m is s io n , p r o v id e d t h a t a c c u r a te acknow ledgm ent o f s o u rc e i s m ade. R e q u e s ts f o r p e r m is s io n f o r e x te n d e d q u o ta t io n from o r r e p r o d u c t io n o f t h i s m a n u s c r ip t i n w ho le o r i n p a r t may b e g r a n te d b y th e h e a d o f th e m a jo r d e p a r tm e n t o r t h e Dean o f t h e G ra d u a te C o l le g e when i n h i s ju d g m en t t h e p ro p o s e d u s e o f t h e m a t e r i a l ife i n t h e i n t e r e s t s o f s c h o la r s h i p . I n a l l o th e r i n s t a n c e s , h o w ev er, p e rm is s io n m u st b e o b ta in e d from t h e a u th o r .
SIGNED:
APPROVAL BY THESIS DIRECTOR
T h is t h e s i s h a s b e e n a p p ro v e d on th e d a te shown b e lo w :
~ A r th u r r . KemmererP r o f e s s o r o f A g r i c u l t u r a l B io c h e m is try
ii
![Page 4: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/4.jpg)
ACKNOWLEDGMENTS
The a u th o r e x p re s s e s s in c e r e g r a t i t u d e :
To D r. E . B. K u r tz , J r . f o r th e t o p i c o f t h i s t h e s i s .
' To D rs . A. R. Kem m erer, K. M a tsu d a , and M. Z a i t l i n f o r
c o u n s e l and g u id a n c e d u r in g th e c o u rs e o f t h i s s tu d y and i n
p r e p a r a t i o n o f t h i s t h e s i s .
iii
![Page 5: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/5.jpg)
TABLE OF CONTENTS
Page
INTRODUCTION .................................................................................................................... 1
REVIEW OF LITERATURE.......................................................... 2
EXPERIMENTAL METHODS................................................................................................. 4
RESULTS AND DISCUSSION ............................................................................................ 18
SUMMARY.............................................................................................................................. 31
LITERATURE C IT E D ...................................... 32
iv
![Page 6: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/6.jpg)
LIST OF TABLES
T a b le Page
I . C om parison o f t o t a l s a l t and io n c o n c e n t r a t io n s o fth e n u t r i e n t m ed ia ( m m o l e s / l i t e r ) ............................................ .... . 11
I I P h o sp h a te c o n te n t o f n u t r i e n t m ed ia (m M )....................................... l 4
v
![Page 7: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/7.jpg)
LIST OF FIGURES
F ig u re Page
1 S ta n d a rd c u rv e f o r p h o s p h a te a n a ly s e s b y th e K i t s o n -M ello n m e t h o d ............................................................................................... 13
2 The f r e s h w e ig h t o f N. c o c h i s e n s i s grow n on v a r io u sam ounts o f p h o s p h a te ............................................................................................. 2b
3 The f r e s h w e ig h t o f N. s i n u a t a grow n on v a r io u sam ounts o f p h o s p h a te . . . . . ................................................................... 25
4 The f r e s h w e ig h t o f N. c o c h i s e n s i s grown on m ed ia c o n ta in in g a d e f i c i e n c y o r an e x c e s s o f p h o s p h a teo r s u l f a t e ......................................................................................................... 26
5 The f r e s h w e ig h t o f N. s i n u a t a grown on m ed ia c o n ta in in ga d e f i c i e n c y o r an e x c e s s o f p h o s p h a te o r s u l f a t e . . . . 27
6 P l a t e s o f N. c o c h i s e n s i s and N. s i n u a t a p r o t h a l l i agrow n on d i f f e r e n t a g a r m e d i a ........................................................... . 28-30
vi
![Page 8: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/8.jpg)
ABSTRACT
I n 1963 i t was r e p o r t e d t h a t N. c o c h is e n s i s w ould grow
e q u a l ly w e l l on b o th K n o p 's and P r a n t l ' s a g a r m ed ia w h i le Eh s in u a t a
w ould grow o n ly on th e fo rm e r medium. S in c e P r a n t l ' s medium la c k s
th e p h o s p h a te io n and c o n ta in s an e x c e s s o f t h e s u l f a t e i o n , i t was
p o s t u l a t e d t h a t N. c o c h i s e n s i s h a s a v e ry low p h o s p h a te r e q u ir e m e n t
o r i s a b le t o s u b s t i t u t e th e s u l f a t e io n f o r th e p h o s p h a te io n .
The p u rp o se o f t h i s t h e s i s was t o show th e lo w e s t l e v e l o f p h o s p h a te
r e q u i r e d by b o th f e r n s and t o show th e i n f lu e n c e o f th e s u l f a t e io n
on th e am ount o f g r o w th .
E x p e r im e n ta l ly , p r o t h a l l i a o f b o th s p e c ie s w ere c u l t u r e d on
a g a r m ed ia t h a t c o n ta in e d v a r io u s am ounts o f p h o s p h a te and s u l f a t e .
E ach a g a r medium was a n a ly z e d f o r p h o s p h a te , and th e g ro w th re s p o n s e s
w ere d e te rm in e d by p h o to g ra p h ic and f r e s h w e ig h t s t u d i e s .
When th e r e s u l t s - o f t h e s e s t u d i e s w ere o b t a in e d , i t was found
t h a t K. s i n u a t a , a s w e l l a s N. c o c h i s e n s i s , w ould grow on P r a n t l ' s
m edium . I n b o th c a s e s th e f e r n s grown on P r a n t l ' s medium h ad a lo w er
f r e s h w e ig h t th a n th o s e grown on K n o p 's m edium. U nder th e e x p e r im e n ta l
c o n d i t io n s u s e d , b o th f e r n s r e q u i r e d 0 .8 mM HPO^ and 2 .7 mM 80^ p lu s t h e
o th e r io n s o f K n o p 's s o l u t i o n f o r m ax im al g ro w th . No p r o t h a l l i a ! g ro w th
was e v id e n t on m ed ia c o n ta in in g no p h o s p h a te and s u l f a t e . B o th f e r n
s p e c ie s showed some g ro w th on m ed ia c o n ta in in g no p h o s p h a te b u t h ig h
am ounts o f s u l f a t e . On th e s e m ed ia b o th f e r n s h ad l a r g e r c e l l s th a n th o s e
vii
![Page 9: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/9.jpg)
viiigrown on m ed ia l a c k in g p h o s p h a te and s u l f a t e . When b o th f e r n s w ere
■grown on a d e q u a te am ounts o f p h o s p h a te , t h e r e was an i n c r e a s e i n
c e l l s i z e and num ber.
![Page 10: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/10.jpg)
INTRODUCTION
In 1959 H ev ly (6 ) r e p o r t e d t h a t th e x e r o p h y t ic ro c k f e r n ,
N o th o la e n a c o c h i s e n s i s G oodd ing , w ould grow e q u a l ly w e l l on b o th K n o p 's
(2 ) and P r a n t l ' s (2 ) m e d ia ; h o w ev er, a s i s t e r s p e c ie s i n t h e N o th o la e n a
co m p lex , N o th o la e n a s i n u a t a (L a g a sc a ex S w a r tz ) K a u l f u s s , w ould d ie i f
grow n on th e l a t t e r m edium . K n o p 's s o l u t i o n i s a b a la n c e d n u t r i e n t
s o l u t i o n w h i le P r a n t l ' s s o l u t i o n l a c k s t h e p h o s p h a te io n and h a s an
e x c e s s o f s u l f a t e i o n ( ( 2 ) . The r e s u l t s o f H e v ly ' s e x p e r im e n t i n d i c a t e
t h a t th e p h o s p h a te r e q u i r e m e n t o f N. c o c h i s e n s i s i s l e s s th a n t h a t o f
N. s i n u a t a . S in c e t h e r e a r e no r e p o r t s i n th e l i t e r a t u r e c o n c e rn in g t h e
p h o s p h a te r e q u i r e m e n t o f t h e s e f e r n s , an e x p e r im e n t was d e s ig n e d t o
d e te rm in e th e lo w e s t l e v e l o f p h o s p h a te io n n e c e s s a r y t o s u s t a i n n o rm a l
g ro w th . I n a d d i t i o n , t h e r e i s a s u g g e s t io n t h a t s u l f a t e may s u b s t i t u t e
p a r t i a l l y f o r p h o s p h a te i n p l a n t s ( 9 ) . A s tu d y a l s o was made to
d e te rm in e i f g ro w th o f f e r n s c o u ld o c c u r on p h o s p h a te d e f i c i e n t b u t
s u l f a t e c o n ta in in g m e d ia .
1
![Page 11: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/11.jpg)
REVIEW OF LITERATURE
The M o th o la e n a f e r n s u se d i n t h i s t h e s i s a r e u s u a l l y found
.grow ing on n o r th o r w e s t - f a c in g s lo p e s o f ro c k l e d g e s , c l i f f s , canyon
w a l l s , b o u l d e r s , o r on t h i n , ro c k y s o i l s . N. c o c h i s e n s i s n o rm a lly
grow s on c a lc a r e o u s s a n d s to n e o r l im e s to n e in w h ich th e pH i s n e u t r a l
o r s l i g h t l y b a s i c w h e re a s N. s i n u a t a grow s on ig n e o u s and m etam o rp h ic
ro c k s su ch a s g n e i s s , q u a r t s i t e , o r g r a n i t e i n w h ich th e pH i s n e u t r a l
o r s l i g h t l y a c id i c ( 5 , 6 ) . I n t h e s e h a b i t a t s , th e f e r n s o f t e n a r e
ex p o sed t o e x c e s s e s o r d e f i c i e n c i e s o f w a te r and v a r io u s m in e r a l s ( 6 ) .
S in c e t h e two f e r n s p e c ie s grow w e l l u n d e r w id e ly d i f f e r e n t and
som etim es h a r s h c o n d i t i o n s , t h e i r n u t r i e n t r e q u i r e m e n ts may be v e ry
d i s s i m i l a r . H e v ly 's s t u d i e s w i th n u t r i e n t m ed ia c u l t u r e s s u g g e s t t h a t
t h i s i s , i n f a c t , th e c a s e ; th e p h o sp h a te r e q u ir e m e n t o f N. c o c h i s e n s i s
a p p e a rs t o b e c o n s id e r a b ly l e s s th a n N. s i n u a t a .
S o lu t io n c u l t u r e te c h n iq u e s h av e b e e n u s e d p r e v i o u s ly t o a s s e s s
t h e n u t r i e n t r e q u i r e m e n ts o f f e r n s . I n 1920 Brown (2 ) r e p o r t e d t h a t
B i e j e r n c k 's , B i m e r and Luc a n u s 's , K n o p 's , P r a n t i ' s , and S a c h 's s o l u t i o n s
w ere f a v o r a b le f o r g ro w th o f th e P o ly p o d ia c e a f e r n s . She a l s o s t a t e d
t h a t K n o p 's and P r a n t i ' s s o l u t i o n s w ere b e s t s u i t e d f o r p r o t h a l l i a g ro w th .
S in c e b o th Brown and H ev ly showed t h a t some f e r n s p e c i e s c o u ld grow on
p h o s p h a te d e f i c i e n t b u t s u l f a t e c o n ta in in g m e d ia , i t a p p e a r s l i k e l y t h a t
some s u l f a t e r e p la c e m e n t o f p h o s p h a te can o c c u r , o r t h e r e a r e p h o s p h a te -
s u l f a t e i n t e r a c t i o n s w h ich a r e n o t u n d e r s to o d .
2
![Page 12: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/12.jpg)
3V a r io u s i n t e r a c t i o n s o f p h o s p h a te and s u l f a t e h av e "been shown
t o e x i s t i n p l a n t s . C o ic , L e s a in t , and G ra n d je a n ( 3 ) fo u n d t h a t a l th o u g h
th e g r a i n and s tr a w y i e l d s w ere l e s s i n p h o s p h a te d e f i c i e n t b a r l e y , th e
u p ta k e o f s u l f a t e was g r e a t e r th a n i n p l a n t s grown on p h o s p h a te c o n ta in in g
n u t r i e n t s o l u t i o n s . A ls o , i n h ig h e r p l a n t s , F u j iw a ra and T o r i i (4 )
showed t h a t when s u l f a t e i s l a c k i n g , t h e am ount o f p h o s p h a te i s in c re a s e d ; .
On New Z e a la n d p a s t u r e s d e f i c i e n t i n s u l f a t e and p h o s p h a te L am erink (1 0 )
fo u n d t h a t th e y i e l d o f c o c k s f o o t c o u ld b e i n c r e a s e d b y th e a d d i t i o n o f
p h o s p h a te o r s u l f a t e , b u t t im o th y y i e l d s . c o u l d b e i n c r e a s e d o n ly w i th
s u l f a t e . By a d d in g s u l f a t e and p h o s p h a te to g e th e r , l a r g e i n t e r a c t i o n s
t h a t in c r e a s e d th e h e rb a g e y i e l d o f w h i te c lo v e r and r y e g r a s s w ere ' . . . ; V . i '
o b s e rv e d . A s i m i l a r e f f e c t o f s u l f a t e and p h o s p h a te was a l s o o b s e rv e d b y
T a k a h a sh i e t a l , ( 1 2 ) ) They fo u n d t h a t t h e two io n s en h a n ce d th e
s t im u la to r y e f f e c t o f g i b b e r e l l i n on g e rm in a t io n o f N. tabacum s e e d s .
In v iew o f t h e p h o s p h a te - s u l f a t e i n t e r a c t i o n s , i t a p p e a r s l i k e l y t h a t
s u l f a t e can p a r t i a l l y r e p l a c e p h o s p h a te i n h ig h e r p l a n t s .
To show t h a t t h i s s u b s t i t u t i o n may e x i s t , W ald (1 3 ) s t a t e s t h a t
s u l f a t e and p h o s p h a te are; s i m i l a r i n t h a t th e y h ave w eak , w id e -s p a c e d -
bonds w h ich a r e v u ln e r a b le t o a t t a c k b y o t h e r m o le c u le s t h a t p rom o te
exchange r e a c t i o n s .
K u rtz and .W aldon ( 9 ) s tu d i e d th e u t i l i z a t i o n o f s u l f a t e i n •
compounds w h ich n o rm a lly c o n ta in p h o s p h a te . U s in g th e o a t p l a n t , th e y
h ave shown t h a t a s u l f a t e d s u g a r ( s ) was fo rm ed when th e o a t was grow n f o r
t h r e e w eeks on a p h o s p h a te d e f i c i e n t m edium . T h e r e f o r e , b e s id e s
p h o s p h o ry la te d s u g a r s , a t l e a s t one s u l f a t e d s u g a r can e x i s t i n h ig h e r p l a n t s .
![Page 13: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/13.jpg)
k
S u l f a t e h a s a l s o b e e n shown t o o c c u r i n h ig h e n e rg y com pounds.
B a n d u r s k i , W ils o n , and S q u ire s ( l ) showed t h a t an enzyme sy s te m e x i s t s ,
i n y e a s t t h a t com bines s u l f a t e w i th ATP^" t o fo rm PAPS. T h is sy s te m was
fo u n d t o c o n ta in two enzym es: ATP s u l f u r a s e , w h ich c a t a ly z e s th e
fo rm a tio n o f APS from ATP and s u l f a t e , and APS k i n a s e , w h ich u s e s ATP
t o c o n v e r t APS t o PAPS. A lth o u g h th e e x i s t e n c e o f h ig h e n e rg y s u l f a t e
compounds h a s b e e n shown i n y e a s t and h ig h e r p l a n t s , s u b s t i t u t i o n f o r
h ig h e n e rg y p h o s p h a te compounds h a s n o t y e t b e e n e s t a b l i s h e d .
I n v ie w o f th e f in d i n g s t h a t s u l f a t e can be u s e d i n p h o s p h o ry la te d
i n te r m e d ia t e s and i n h ig h e n e rg y com pounds, one i s l e d t o s p e c u la te
t h a t s u l f a t e may p a r t i a l l y s u b s t i t u t e f o r t h e p h o s p h a te r e q u i r e m e n t i n
some o rg a n is m s . T h e r e f o r e , i n d e s ig n in g m ed ia 1 t o 6 t o d e te rm in e th e
lo w e s t p h o s p h a te l e v e l r e q u i r e d b y th e f e r n s , am ounts o f KgHPO^ w ere
o m it te d and e q u a l am ounts o f KgSO^ w ere ad d ed t o k eep th e am ount o f K
c o n s ta n t and t o show th e in f lu e n c e o f s u l f a t e .
EXPERIMENTAL METHODS
The m ed ia u s e d i n t h i s s tu d y w ere d e s ig n e d t o d e te rm in e th e lo w e s t
p h o s p h a te l e v e l r e q u i r e d b y th e . f e r n s and t o show th e i n f l u e n c e o f s u l f a t e .
L arg e q u a n t i t i e s o f n u t r i e n t a g a r m ed ia w ere p r e p a r e d , and 10 m l a l i q u o t s
c o n ta in in g 0 .2 g a g a r w ere t r a n s f e r r e d t o 2 x J r 'in c h p e t r i d i s h e s .
A l l s o l u t i o n s e x c e p t Ho. 11 c o n ta in e d , i n e a c h l i t e r , 2 m l o f K u r t z 's m in o r
**■ A b b r e v ia t io n s u s e d a r e : PAPS, a d e n o s in e - 3 ' - p h o s p h a te - 5 ' -p h o s p h o s u l f a te ; APS, a d e n o s in e - 5 ' - p h o s p h o s u l f a t e ; P ^ , o r th o p h o s p h a te ;ATP, a d e n o s in e t r i p h o s p h a t e .
![Page 14: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/14.jpg)
The pH v a lu e s o f e a che le m e n t s o l u t i o n (8 ) and 1 d ro p o f Vj0 F e C lg .
s o l u t i o n w ere a d ju s t e d t o 7 .5 w i th 2N NaOH. E le v e n d i f f e r e n t s o l u t i o n s
w ere u s e d , and th e c o m p o s i t io n s a r e th e f o l l o w in g :
S o lu t io n X (K h o p 's s o l u t i o n )
S a l t C o n e e n t r a t i on o f S a l t
g / l i t e r mM
MgSO^ 0 .2 5 2 .1
CafNOgjg.ltHgO 1 .0 0 4 .2
KC1 0 .1 2 1 .6
KgHPO^ 0 .2 5 1 .4
0 ,6 5 m l 2 N HaOH 1 .2
T o ta l 1 0 .5
S o lu t io n 2
S a l t C o n c e n t r a t io n o f S a l t
g / l i t e r mM
MgS04 0 .2 5 2 .1
c a ( m 3 )2 .4 p 2o 1 .0 0 4 .2
KC1 0 .1 2 1 .6
K2HP04 0 .2 0 1 .2
KgSC^ 0 .0 5 0 .3
0 .5 0 m l 2 H HaOH 1 .0
T o ta l i o . 4
![Page 15: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/15.jpg)
Solution 3
' S a l t C o n c e n t r a t io n o f S a l t
g / l i t e r mM
MgSO^ 0 .2 5 2 . 1
Ca(H03 )2 .UH20 1 .00 4 .2
KC1 0 .12 1 . 6
KgHPO^ 0 .1 5 0 . 8
K2S0i+ 0 .10 0 . 6
0 .4 0 m l 2 N MaOH 0 . 8
T o ta l 1 0 .1
S o lu t io n 14-
S a l t C o n c e n t r a t io n o f S a l t
g / l i t e r mM
MgSO^ 0 .2 5 2 . 1
Ca(N03 ) 2 .4H20 1 .0 0 4 .2
KC1 0 .12 1 . 6
KgHPO^ 0 ,1 0 0 . 6
KgSO^ 0 .1 5 0 . 8
0 .3 0 m l 2 N NaOH 0 . 6
T o ta l 9 -9
![Page 16: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/16.jpg)
Solution 5
S a l t C o n c e n t r a t io n o f S a l t
g / l i t e r mM
MgS04 0 .2 5 2 .1
Ca($r03 )2 .4H20 1 ,0 0 4 .2
KC1 0 .1 2 1 .6
KgHPO^ 0 .0 5 0 .3
k2 so4 0 .2 0 1 .2
0 .2 0 m l 2 N NaOH 0 . 4
T o ta l
S o lu t io n 6
9 . 8
S a l t C o n c e n t r a t io n o f S a l t
g / l i t e r mM
MgSO^ 0 .2 5 2 .1
Ca(N03 ) 2 .4H20 1 .0 0 4 .2
KC1 0 ,1 2 1 .6
K2S04 0 .2 5 1 .4
0 .1 0 m l 2 H NaOH 0 .2
T o ta l 9 -5
![Page 17: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/17.jpg)
S o lu t io n 7 ( P r a n t i ' s s o l u t i o n )
S a l t C o n c e n t r a t io n o f S a l t
g / l i t e r mM
MgSO^ 0 .5 0 4 .2
V O ** 0 .7 0 4 .0
NaCl 0 .2 3 4 .0
CaSO^.PHgO 0 .70 4 .0
0 .0 1 0 . 2
0 .05 m l 2 N NaOH 0 . 1
T o ta l 16 .5
S o lu t io n 8
S a l t C o n c e n t r a t io n o f S a l t
g / l i t e r mM
MgS04 0 .2 5 2 .1
Ca(N03 )2 .4H20 1 .0 0 4 .2
KC1 0 .1 2 1 .6
KgKPO^ 0 .2 5 1 .4
NagSO^ 0 .1 7 . 1 .2
0 .6 0 m l 2 N HaOH 1 .1
T o ta l 1 1 .6
![Page 18: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/18.jpg)
Solution 9
S a l t
MgS04
Ca(N03)2 .4H20
KC1 ■
KgHPO^
KgSOj^
0 .7 5 m l 2 N WaOH
T o ta l
S a l t '
MgClg.SHgO
Ca(N03 )2 .4H20
KC1
0 .1 0 m l 2 N NaOH
T o ta l
C o n c e n t r a t io n o f S a l t
g / l i t e r
0 .2 5
1.00
0 .1 2
0 .2 5
0 .2 5
mM
2.14 .2
1 .6
1 .4
1 .4
1 .5
12.2
S o lu t io n 10
C o n c e n t r a t io n o f S a l t
g / l i t e r
0.68
1.000.22
mM
3 .4
4 .2
2 .9
0.1
10.6
S o lu t io n 11
One l i t e r d e io n iz e d w a te r and 0 .1 m l 2 N HaOH
![Page 19: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/19.jpg)
10T a b le 1 shows a c o m p a riso n o f t h e t o t a l s a l t and io n c o n c e n t r a t io n s
o f th e n u t r i e n t m e d ia . . Medium 1 i s K n o p 's s o lu t i o n p lu s a g a r and i s
u s e d a s t h e f i r s t medium i n a c o n c e n t r a t io n s e r i e s t h a t h a s v a ry in g
am ounts o f p h o s p h a te . S in c e H ev ly (6 ) r e p o r t e d t h a t b o th f e r n s grew
w e l l on K nop’ s m edium , i t s p h o s p h a te c o n c e n t r a t i o n , 1 .4 mM, was th e
maximum u s e d in t h i s s tu d y .
M edia 2 t o 6 r e p r e s e n t th e o t h e r m ed ia i n th e c o n c e n t r a t io n s e r i e s
o f v a ry in g am ounts o f p h o s p h a te ; th e y w ere u s e d t o d e te rm in e th e lo w e s t
l e v e l o f p h o s p h a te r e q u i r e d f o r maximum g ro w th . In t h e i r p r e p a r a t i o n th e
am ounts o f KgHPO^ and KgSO^ w ere r e g u l a t e d t o m a in ta in a c o n s ta n t
p o ta s s iu m c o n c e n t r a t i o n , and th e pH v a lu e s w ere a d ju s t e d t o pH 7 * 5 .
As shown i n T a b le 1 , t h e p h o s p h a te c o n c e n t r a t io n was u l t i m a t e l y lo w e re d
t o 0 .0 mM i n s o l u t i o n 6 , b u t th e s u l f a t e c o n c e n t r a t io n was i n c r e a s e d t o
3 -5 mM.
Medium 7 i s P r a n t i ’ s s o l u t i o n p lu s a g a r . ' T h is medium was p r e p a r e d
t o c o n f irm H e v ly 1s s ta te m e n t ( 5 , 6 ) t h a t N. s in u a t a d i e s and N. c o c h i s e n s i s
l i v e s when grown on i t . I t d i f f e r s from m ed ia 1 t o 6 i n t h a t i t h a s no
p h o s p h a te , b u t i t s s u l f a t e c o n te n t i s a v a lu e 3 t o 4 t im e s h ig h e r th a n i n
t h e fo rm e r s o l u t i o n s .
I n t h i s e x p e r im e n t m ed ia 8', 9? 1 0 , and 11 w ere u s e d a s c o n t r o l
m e d ia . M edia 8 and 9 c o n ta in 3 .5 mM s u l f a t e w i th e x c e s s Na and K.
By a d d in g HagSO^ and KgSO^ t o medium 1 , t h e s e m ed ia w ere p r e p a r e d t o
show t h a t a n y 'p o s s i b l e ch a n g es i n th e am ount o f g ro w th on m ed ia 1 t o 6
was n o t due t o a h a rm fu l e f f e c t o f i n c r e a s i n g am ounts o f s u l f a t e .
![Page 20: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/20.jpg)
11
T a b le 1
C om parison o f T o ta l S a l t and Io n C o n c e n t r a t io n s o f th e N u t r i e n t M edia ( m m o le s / l i t e r )
S o l .No. S a l t Na K Ca Mg N°3 HP04 s o u C l
1 1 0 .5 1 .2 4 .4 4 .2 2 .1 8 .4 1 .4 2 .1 1 .6
2 1 0 .4 1 .0 4 .6 4 .2 2 .1 8 .4 1 .2 2 .4 1 .6
3 1 0 .1 0 .8 4 .4 4 .2 2 .1 8 .4 0 .8 2 .7 1 .6
k 9 -9 0 .6 4 .4 4 .2 2 .1 8 .4 0 .6 2 .9 1 .6
5 9 .8 0 .4 4 .6 4 .2 2 .1 8 .4 0 .3 3 .3 1 .6
6 9 -5 0 .2 4 .4 4 .2 2 .1 8 .4 0 .0 3 .5 1 .6
7 1 6 .5 4 .1 8 .0 4 .0 4 .2 0 .2 0 .0 1 2 .2 4 .0
8 1 1 .6 3 .5 4 .4 4 .2 2 .1 8 .4 1 .4 3 .5 1 .6
9 1 2 .2 1 .5 7 .2 4 .2 2 .1 8 .4 1 .4 3 .5 1 .6
10 1 0 .6 0 .1 2 . 9 4 .2 3 .4 ' 8 .4 0 .0 0 .0 9 -7
11 0 .1 0 .1
![Page 21: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/21.jpg)
12.
Medium 10 was u se d t o d e m o n s tra te g ro w th r e s p o n s e s when b o th s u l f a t e and
p h o s p h a te a r e a b s e n t . S i m i l a r l y , medium 11 was u se d t o show g ro w th
r e s p o n s e s on a g a r when no n u t r i e n t s w ere a d d e d .
U s in g th e m ethod o f K its o n and M ello n ( ? ) , p h o s p h a te a n a ly s e s
w ere p e rfo rm e d on th e m ed ia and th e s p o r e s . T h is m ethod was s e l e c t e d
b e c a u s e th e s e n s i t i v i t y i s 1 ppm, t h e c o lo r r e a c t i o n i s s t a b l e , and
i n t e r f e r e n c e from o th e r d o n s i n th e medium i s n e g l i g i b l e . To p r e p a r e a
n u t r i e n t medium f o r a n a l y s i s , a 25 m l sam ple o f t h e n u t r i e n t s o l u t i o n
c o n ta in in g a g a r was p i p e t t e d i n t o a c r u c i b l e . Ten r e p l i c a t e a n a ly s e s
w ere ru n from e a c h n u t r i e n t m edium ; A f te r c a r e f u l l y d r y in g th e w a te r
from th e sam p les i n t h e c r u c i b l e s b y p l a c i n g them in a d ry in g o v e n , th e
sam p les w ere i g n i t e d and a sh e d a t 5 0 0 ° . The c o n te n t s o f t h e c r u c i b l e s
w ere ta k e n up in 10 m l o f 1 N HKO^ and q u a n t i t a t i v e l y t r a n s f e r r e d t o
100 m l v o lu m e tr ic f l a s k s . The f l a s k s w ere d i l u t e d t o v o lu m e , and 2 5 -35
m l a l i q u o t s c o n ta in in g 10 -2 0 0 jig HPOj ̂ w ere t r a n s f e r r e d t o 50 m l v o lu m e tr ic
f l a s k s . F iv e m i l l i t e r s ea ch o f 7 .5 N HNOg, 0 .0 2 M HH^VOg, and 0 .0 7 M
(nH^)^MQyOg^. 4Hg0 w ere added i n o r d e r , and th e f l a s k s w ere b ro u g h t t o
volum e w i th d e io n iz e d w a te r and s h a k e n . A f t e r o n e - h a l f h o u r , th e
a b s o rb a n c e o f th e sam ple was d e te rm in e d w i th a Beckman DU s p e c t r o p h o to m e te r
a t th e w a v e le n g th o f maximum a b s o r p t io n , 3&5 n y - The r e s u l t s w ere com pared
t o s ta n d a r d p h o s p h a te s o l u t i o n s , p r e p a r e d and a n a ly z e d a t th e same t im e .
An exam ple o f t h e s ta n d a r d p h o s p h a te c u rv e i s shown i n F ig u re 1 ,
The r e s u l t s o f th e p h o s p h a te a n a ly s e s on th e m ed ia c o n ta in in g a g a r
a r e shown i n T a b le 2 . I n a l l c a s e s th e p h o s p h a te fo u n d b y a n a ly s e s was
n e a r l y e q u a l t o th e t h e o r e t i c a l am ount added b y th e n u t r i e n t s o lu t i o n a lo n e
![Page 22: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/22.jpg)
OPT
ICA
L D
EN
SIT
Y
.900-1
.8 0 0 -
.7 0 0 -
,6 0 0 -
.5 0 0 -
4 0 0 -
3 0 0 -
PHOSPHATE— mmoles in reoction mixture
F i g u r e 1 . S t o n d a r d C u r v e f o r P h o s p h a t e A n a l y s e s by t h e K i t s o n - M e I I on M e t h o d .
HU>
![Page 23: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/23.jpg)
Table 2P h o sp h a te C o n te n t o f N u t r i e n t M edia8, (mM)
' M edia No. T h e o r e t i c a l i n medium
(mM)
A n a ly se s o f sam p les^
(mM)
• 1 1 .4 3 1 .4 2
2 1 .1 5 l . l 6
3 0 .86 0 .8 9
4 0 .5 8 0 .52
5 0 .2 9 0 .2 7
6 0 .0 0 0 .05
7 0 .0 0 0 .03
8 1 .4 4 1 .5 5
9 1 .4 4 1 . 4 i
10 0 .0 0 o .o 4
i i 0 .0 0 0 .0 2
a N u t r i e n t m ed ia i n d i c a t e s n u t r i e n t s o l u t i o n p lu s a g a r ,
k I n d i c a t e s a v e ra g e am ount o f 10 r e p l i c a t e s on th e medium.
![Page 24: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/24.jpg)
15F o r exam ple, 1 .4 3 mmoles HPO^ was added t o ea ch l i t e r o f medium 1 ; th e
a v e ra g e am ount fo u n d was 1 .4 2 mM. As shown i n T a b le 2 , m ed ia 6 , 7 , 1 0 ,
and 11 c o n ta in e d no ad d ed p h o s p h a te , b u t s m a l l am ounts w ere fo u n d .
F o r e x a m p le , i n medium 6 , 0 ,0 4 mM was fo u n d . T h is s m a l l am ount o f
p h o s p h a te can p r o b a b ly be a t t r i b u t e d t o e r r o r s i n a n a l y t i c a l p r o c e d u r e s .
S in c e th e s p o re s th e m s e lv e s may h av e h ig h l e v e l s o f endogenous
p h o s p h o ru s , t h e i r c o n t r i b u t i o n t o t h e medium was a l s o d e te rm in e d .
When 4 5 .8 mg o f N. c o c h i s e n s i s s p o r e s , r e p r e s e n t i n g a p p ro x im a te ly 2800
s p o r e s , w ere a n a ly z e d f o r p h o s p h a te , 0 .233 mmoles HPO^ was fo u n d .
By u s in g 58 a s t h e a v e ra g e num ber o f g e rm in a t in g s p o re s p r e s e n t i n e a c h
d i s h , i t was c a l c u l a t e d t h a t 4 .1 x 10*"3 mM HPO^ was c o n t r i b u t e d b y th e
s p o r e s . T h e r e f o r e , t h e am ount o f p h o s p h a te c o n t r i b u t e d b y N. c o c h i s e n s i s
i s i n s i g n i f i c a n t .
F o llo w in g a m ethod d e s c r ib e d b y H evly ( 6 ) , s p o re s o f b o th
N o th o la e n a s p e c i e s w ere o b ta in e d from f e r t i l e f ro n d s t h a t w ere c o l l e c t e d
i n th e T ucson a r e a . The s p o re s w ere p la c e d i n v i a l s and s to r e d i n th e
d a rk t o p r e v e n t p re m a tu re g e m i n a t i o n ( 5 , 6 ) . The f e r n s w ere i d e n t i f i e d
b y D r. H ev ly and a r e on f i l e a t th e U n i v e r s i t y o f A r iz o n a H e rb a riu m .
A m ethod o f s p o re s t e r i l i z a t i o n ( l 4 ) was o b ta in e d from th e
l i t e r a t u r e and m o d if ie d . By u s in g d i s p o s a b le t r a n s f e r p i p e t t e s ,
a p p ro x im a te ly 100 mg o f s p o re s w ere p la c e d on g l a s s w ool i n t h e b o tto m
o f a 7 x ■§- in c h c h ro m a to g ra p h ic colum n t h a t h ad b e e n s t e r i l i z e d p r e v i o u s l y .
The g l a s s w ool a llo w e d l i q u i d s t o d r a i n b u t d id n o t a l lo w th e s p o re s t o
p a s s th r o u g h . To p r e v e n t c lu m p in g , th e s p o re s w ere t r e a t e d b y im m ers in g
th e colum n i n s t e r i l e 0 .1 % .Tween 80 f o r 20 m in u te s . T h is w as fo llo w e d
![Page 25: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/25.jpg)
16
by e x p o s in g th e s p o re s t o 30$ Chlorox"*" f o r 20 m in u te s . The s p o re s
w ere th e n r i n s e d once w i th s t e r i l e , d e i o n i z e d w a te r and f i n a l l y su sp e n d e d
i n 35 m l o f s t e r i l e , d e io n iz e d w a te r . S t e r i l e p e t r i d i s h e s , c o n ta in in g
d i f f e r e n t n u t r i e n t a g a r m e d ia , w ere i n o c u la te d w i th 0 .3 m l o f th e s p o re
s u s p e n s io n s . T h is gave ah a v e ra g e num ber o f 58 and 65 g e rm in a t in g
s p o re s f o r N. c o c h i s e n s i s and N. s i n u a t a , r e s p e c t i v e l y . A f t e r i n o c u l a t i o n
th e p e t r i d i s h e s w ere k e p t i n a c u l t u r e room , m a in ta in e d a t 30° , and
r e c e iv e d c o n tin u o u s l i g h t i n t e n s i t i e s o f 700 -900 f t - c from b an k s o f
c o o l w h i t e , f l u o r e s c e n t l i g h t s . The l i g h t i n t e n s i t i e s w ere m easu red
w i th a W eston 603 p h o to m e te r . S t e r i l e , d e io n iz e d w a te r ( 0 .2 - 0 .3 ) m l ) was
added a t l e a s t once a .w eek t o m a in ta in th e m o is tu re c o n te n t o f th e a g a r
m e d ia . S in c e t h e l i g h t i n t e n s i t i e s v a r i e d from 700 -900 f t - c on th e
g ro w in g t a b l e , t h e d i s h e s w ere r e a r r a n g e d a f t e r ea ch a d d i t i o n o f w a t e r .
The f r e s h w e ig h t o f t h e p r o t h a l l i a l c o n te n t s o f e a c h d i s h was
d e te rm in e d by f i r s t lo o s e n in g th e p r o t h a l l i a from th e a g a r w i th w a te r .
When th e p r o t h a l l i a w ere f r e e d from th e a g a r , th e y w ere c o l l e c t e d on 4 cm
Whatman N o .,1 f i l t e r p a p e r i n a s m a ll B uchner f u n n e l , rem oved w ith a
s p a t u l a o r tw e e z e r s , and p la c e d in a s m a l l , t a r e d v i a l . The s u r f a c e o f
t h e a g a r m ed ia and f i l t e r p a p e r s w ere ch e ck e d w i th a d i s s e c t i n g m ic ro sc o p e
t o be s u re t h a t a l l p r o t h a l l i a and t h e i r r h i z o i d s w ere added t o t h e v i a l .
I f th e p r o t h a l l i a o r th e v i a l h ad d ro p s o f w a te r on t h e i r s u r f a c e , th e
f r e s h w e ig h t was n o t m e a su re d . Once th e i n i t i a l p r o t h a l l i a l w e ig h ts
1 C h lo ro x i s t h e co m m erc ia l name f o r 5 .2 5 $ NaOCl (by w e ig h t ) .
![Page 26: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/26.jpg)
17w ere o b ta in e d , f r e s h w e ig h t m easu rem en ts w ere made e v e ry 5 days u n t i l
t h e t h i r t y - f i f t h d ay a f t e r g e r m in a t io n .
The m o rp h o lo g ic a l c h an g es w h ich o c c u r r e d d u r in g g ro w th w ere
r e c o r d e d on 35 mm s l i d e p i c t u r e s , ta k e n th ro u g h a 10X l e n s o f a Z e is s
208359^ GFL m ic ro s c o p e . The p i c t u r e s w ere ta k e n on Kodachrom e I I c o lo r
s l i d e f i l m , u s in g a Z e is s Ik o n cam era s e t a t l / 2 5 o f a s e c o n d . A Z e is s
LTKL Nr t r a n s f o r m e r s e t a t 8 was u s e d t o c o n t r o l t h e l i g h t s o u r c e , and
th e e m i t te d l i g h t was f i l t e r e d w ith a b lu e f i l t e r .
![Page 27: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/27.jpg)
RESULTS AM) DISCUSSION
To a v o id m ic r o b ia l g ro w th , t h e a g a r was a u to c l a v e d , and th e
s p o re s w ere s t e r i l i z e d w i th a d i s i n f e c t a n t p r i o r t o g e r m in a t io n .
A 20 m in u te t r e a tm e n t w i th 30% C h lo ro x was fo u n d t o be th e maximum
a l lo w a b le t r e a tm e n t o f th e s p o r e s . The t r e a tm e n t rem oved m ost o f th e
m o ld , b a c t e r i a l , and a l g a l c o n ta m in a t io n from th e s p o re s u r f a c e b u t
d id n o t e l i m i n a te t h e im p e r f e c t fu n g i A l t e r n a r i a , D i p l o d ia , and
S p o ro tr ic h u m . The C h lo ro x t r e a tm e n t a l s o h ad a h a rm fu l e f f e c t on
th e s p o re s s in c e some d id n o t g e r m in a te . I n a d d i t i o n , t h e am ount o f
g ro w th o f th e p r o t h a l l i a was l e s s th a n when s p o re s w ere g iv e n s h o r t e r
e x p o s u re s t o C h lo ro x .
When th e s p o re s o f b o th N o th o la e n a s p e c ie s w ere p la c e d on
n u t r i e n t m ed ia and e x p o sed t o c o n tin u o u s l i g h t i n t e n s i t i e s o f 700 t o
900 f t - c , g e rm in a t io n o c c u r r e d on th e s i x t h day a f t e r i n o c u l a t i o n .
I n b o th s p e c i e s th e m o rp h o lo g ic a l ch a n g es w h ich o c c u r r e d a f t e r g e rm in a t io n
w ere s i m i l a r t o th o s e o f t h e b ra c k e n f e r n P te r id iu m a q u il in u m ( l l ) .
A f t e r im b ib in g w a te r , th e s p o rb s r u p tu r e d , and a g r e e n , u n s e p ta t e
f i l a m e n t and one r h i z o i d em erged . W ith in a few d ay s t h e f i l a m e n t
becam e s e p t a t e s and o t h e r r h i z o i d s form ed on i t . I s o l a t e d p r o t h a l l i a
w ould u n d e rg o f u r t h e r d e v e lo p m en t b y p ro d u c in g a p l a t e l e t from th e
f i l a m e n t . T h ese p l a t e l e t s c o n tin u e d t o d e v e lo p i n an i r r e g u l a r sh ap e
b e c a u s e o f c o n t in u o u s , h ig h l i g h t i n t e n s i t i e s ( 9 , 6 ) . When th e
p r o t h a l l i a w ere c ro w d ed , t h e f i l a m e n ts d id n o t d e v e lo p i n t o p l a t e l e t s .
18
![Page 28: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/28.jpg)
19The r e s u l t s o f f r e s h w e ig h t s tu d i e s showed t h a t N. c o c h i s e n s i s
and N. s i n u a t a re s p o n d e d s i m i l a r l y t o d i f f e r e n t n u t r i e n t m e d ia . T h is i s
shown i n F ig u r e s 2 , 3s ^s and 5 . F o r b o th s p e c ie s a f i f t e e n day p e r io d
a f t e r t h e •o n s e t o f g e rm in a t io n was r e q u i r e d b e f o r e p r o t h a l l i a l w e ig h ts
c o u ld be o b ta in e d on m ed ia 1 , 2 , 3s 4 , 7 S 8 , and 9- W ith th e e x c e p t io n
o f medium 7 , P r a n t i ' s m edium , t h e s e m ed ia had a t l e a s t 0 .6 mM p h o s p h a te
and 2 .1 mM s u l f a t e . I n c o n t r a s t a 20 d ay p e r io d was n e e d ed on m ed ia
5 and 6 , w h ich c o n ta in e d 3 -3 and 3*5 mM s u l f a t e and 0 .3 and 0 .0 mM
p h o s p h a te , r e s p e c t i v e l y , b e f o r e e i t h e r s p e c ie s y i e l d e d s u f f i c i e n t
p r o t h a l l i a f o r w e ig h in g . On m ed ia 10 w h ich h a s no s u l f a t e o r p h o s p h a te
and m ed ia 11 w h ich h a s no n u t r i e n t s , t h e p r o t h a l l i a l w e ig h ts w ere
i n s u f f i c i e n t f o r d e te r m in a t io n even a f t e r g ro w in g 35 d a y s .
The r e s u l t s from m ed ia 1 and 7 d i f f e r i n two ways from th o s e o f
H evly ( 6 ) , who r e p o r t e d t h a t W. c o c h i s e n s i s w ould grow e q u a l ly w e l l on
b o th K hop' s and P r a n t l ' s a g a r medium and N. s i n u a t a w ould n o t grow on
P r a n t i ' s m edium . F i r s t , N. c o c h i s e n s i s d id n o t grow e q u a l ly w e l l on
K n o p 's and P r a n t l ' s m e d ia , a n d , s e c o n d ly , N. s i n u a t a d id grow when '
p l a c e d on P r a n t l ' s m edium .
When s p o re s w ere p la c e d on m ed ia 10 and 1 1 , t h e p r o t h a l l i a
g e rm in a te d a t th e same tim e a s th o s e grown on m ore b a la n c e d m e d ia .
The am ount o f g ro w th on medium 11 shows t h a t c e r t a i n n u t r i e n t s r e q u i r e d
f o r n o rm a l g ro w th a r e a b s e n t i n a g a r a lo n e w h ile th e am ount o f g ro w th
on medium 10 shows t h a t p h o s p h a te and s u l f a t e c a n n o t b o th be e x c lu d e d
from th e a g a r medium.
![Page 29: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/29.jpg)
20F ig u r e s 2 and 3 show t h a t N. c o c h i s e n s i s and N. s i n u a t a grew
s i m i l a r l y on n u t r i e n t m ed ia 1 t o 3 . Medium 1 , K n o p 's m edium , i s a
b a la n c e d n u t r i e n t medium w i th p h o s p h a te and s u l f a t e c o n te n t s o f 1 .4 mM
and 2 .1 mM, r e s p e c t i v e l y . Medium 3 i s s i m i l a r t o medium 1 e x c e p t t h a t
th e p h o s p h a te and s u l f a t e c o n c e n t r a t io n s a r e 0 . 8 and 2 . 7 m&% r e s p e c t i v e l y .
The f a c t t h a t th e g ro w th o f p r o t h a l l i a was a p p ro x im a te ly th e same on
t h e s e m ed ia s u g g e s ts th e l e v e l o f p h o s p h a te i n medium 3 i s n o t l i m i t i n g
g ro w th in t h i s s tu d y .
F ig u r e s 2 and 3 a l s o show t h a t th e am ount o f g ro w th o f b o th f e r n s
on medium 4 was l e s s th a n on m ed ia 1 t o 3- I n t h i s medium th e p h o s p h a te
and s u l f a t e c o n te n t s w ere 0 .6 mM and 2 .9 mM, and th e am ounts o f o t h e r
io n s w ere n o t v a r i e d from th o s e on medium 1 . S in c e a p p ro x im a te ly th e
same num ber o f s p o re s o f e a c h s p e c i e s w ere u s e d p e r d i s h , i t may be
c o n c lu d e d t h a t t h e r e q u i r e m e n ts f o r p h o s p h a te o f th e two s p e c ie s a r e t h e
sam e. The f a c t t h a t g ro w th on medium 4 was l e s s th a n on medium 3 shows
t h a t s u l f a t e c a n n o t s t o i c h i o m e t r i c a l l y r e p l a c e p h o s p h a te . Medium 4
c o n ta in s 0 .6 mM p h o s p h a te and 2 .9 mM s u l f a t e w h e re as medium 3 c o n ta in s .
0 .8 mM HPO4 and 2 .7 mM SO4 .
F ig u r e s 2 and 3 a l s o show t h a t t h e am ounts o f p r o t h a l l i a l g ro w th
on m ed ia 5 and 6 a r e ev en l e s s th a n on medium 4 and f a r l e s s th a n on
medium 1 . A g a in , b o th s p e c ie s r e s p o n d e d s i m i l a r l y . M edia 5 and 6 d i f f e r
from medium 1 b y h a v in g 0 .3 and 0 .0 mM p h o s p h a te , r e s p e c t i v e l y , i n s t e a d
o f 1 .4 mM p h o s p h i t e ; th e y a l s o d i f f e r b y h a v in g 3 -3 and 3 .5 mM s u l f a t e ,
r e s p e c t i v e l y , i n s t e a d o f 2 .1 mM s u l f a t e . From th e am ount o f g ro w th
o b ta in e d on m ed ia 5 and 6 , i t may b e c o n c lu d e d a g a in t h a t s u l f a t e c a n n o t
![Page 30: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/30.jpg)
21
s t o i c h i o m e t r i c a l l y r e p l a c e p h o s p h a te . H ow ever, t h e r e was some g ro w th
on th e s e m ed ia w h e re as no m e a s u ra b le g ro w th was fo u n d u s in g medium 10
w h ich h ad a l l o t h e r io n s e x c e p t p h o s p h a te and s u l f a t e . S in c e m ed ia 5
and 6 d i f f e r from medium 10 p r i m a r i l y i n t h a t th e y c o n ta in d i f f e r e n t
am ounts o f s u l f a t e f o r g ro w th , t h e r e may be a p a r t i a l r e p la c e m e n t o f .
p h o s p h a te b y s u l f a t e .
F ig u r e s 4 and 5, show b o th f e r n s re s p o n d e d q u i t e w e l l t o medium
7 , P r a n t l ' s m edium . T h is medium d i f f e r s from m ed ia 1 t o 6 i n t h a t
o th e r i o n s , i n a d d i t i o n t o p h o s p h a te and s u l f a t e , w ere v a r i e d . I t i s
somewhat s i m i l a r t o th e c o m p o s i t io n o f medium 6 i n t h a t i t h a s no
p h o s p h a te , b u t t h e s u l f a t e c o n te n t i s c o n s id e r a b ly h ig h e r th a n in th e
l a t t e r m edium. When th e am ount o f g ro w th on P r a n t l ' s medium i s com pared
t o t h a t o f medium 1 0 , t h e c o n c lu s io n i s s i m i l a r t o t h a t o b ta in e d when
m ed ia 6 and 10 a r e com pared : th e a d d i t i o n o f s u l f a t e c a u s e s a l a r g e •
i n c r e a s e i n t h e am ount o f g ro w th .
When th e s p o re s o f b o th f e r n s w ere p la c e d on m e d ia 8 and 9 ,
th e am ount o f g ro w th was e q u a l t o t h a t o b ta in e d from medium 1 , a s shown
i n F ig u r e s 4 and 5 . Medium 8 c o n ta in s t h e same io n c o n te n t a s medium 1
e x c e p t t h a t th e Na c o n te n t i s 3 -5 mM i n s t e a d o f 1 .2 mM and th e s u l f a t e
c o n te n t i s 3 •5 mM i n s t e a d o f 2 .1 mM. . S in c e t h e am ounts o f g ro w th on
medium 8 and 9 w ere th e same a s on medium 1 , e x c e s s s u l f a t e i s n o t h a rm fu l
i n th e p r e s e n c e o f e x c e s s Na o r K. T hese r e s u l t s a l s o show t h a t th e
d e c re a s e i n am ounts o f g ro w th on m ed ia 4 , 6 , and 7 i s due t o o m is s io n
o f am ounts o f p h o s p h a te i n t h e medium an d n o t due t o ch a n g es i n th e Na
o r s u l f a t e c o n te n t s .
![Page 31: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/31.jpg)
22The m o rp h o lo g ic a l c h an g es w h ich o c c u r r e d d u r in g g ro w th a r e
shown in F ig u re 6 . P l a t e 1 shows a N. c o c h is e n s i s p r o th a l l i u m on
medium 1 t w e n ty - f iv e d ay s a f t e r g e r m in a t io n . A f te r c a r e f u l o b s e r v a t io n
w i th a d i s s e c t i n g m ic ro s c o p e , p r o t h a l l i a from m ed ia 2 , 3 , 8 , and 9
w ere fo u n d t o be i n d i s t i n g u i s h a b l e from th o s e on medium 1 . The p r o t h a l l i a
a p p e a r a s b ro a d p l a t e l e t s w i th many r h i z o i d s .
P l a t e s 2 and 3 o f F ig u re 6 show p r o t h a l l i a o f N. c o c h i s e n s i s and
H. s i n u a t a , r e s p e c t i v e l y , on medium 4 tw e n ty - f iv e days a f t e r g e rm in a tio n ..
A lth o u g h th e p l a t e s do n o t show e q u a l g ro w th o f b o th s p e c i e s , m ic ro s c o p ic
e x a m in a tio n r e v e a le d t h a t H. c o c h i s e n s i s was n o t d i s t i n g u i s h a b l e fromY , “ ------------------ ■
N. s in u a t a when t h e i r p r o t h a l l i a w ere com pared . P r o t h a l l i a o f b o th f e r n s
grown on medium 4 , w h ic h ..,c o n ta in s 0 .6 mM HPO4 , h ad a b o u t t h e same c e l l
num ber a s th o s e grow n on medium 1 , b u t t h e c e l l s w ere s m a l le r .
T h is s u g g e s ts t h a t c e l l e n la rg e m e n t r a t h e r th a n c e l l d i v i s i o n was h in d e r e d
b y th e low p h o s p h a te c o n te n t .
P l a t e 4 o f F ig u re 6 shows N. c o c h i s e n s i s p r o t h a l l i a on medium 7 ,
P r a n t l ' s m e d iu m ,tw e n ty - f iv e days a f t e r g e r m in a t io n . The p l a t e l e t s a r e
n o t q u i t e a s w ide a s th o s e on m ed ia 1 and 4 , and th e c e l l num ber i s less../;
th a n th o s e o f p r o t h a l l i a grow n on m ed ia 1 and 4 . The c e l l s i z e o f
p r o t h a l l i a on medium 7 i s s i m i l a r t o th o s e o f p r o t h a l l i a from medium 4 .
S in c e t h e f r e s h w e ig h t s tu d i e s showed t h a t th e d e c r e a s e i n am ount o f
g ro w th on m e d ia .4 and 7 was due t o o m is s io n o f p h o s p h a te , i t may b e
c o n c lu d e d t h a t a d e q u a te am ounts o f p h o s p h a te a l s o i n c r e a s e t h e num ber o f
c e l l s i n a p r o th a l l i u m .
![Page 32: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/32.jpg)
23As shown i n P l a t e 5 o f F ig u re 6 , N. c o c h i s e n s i s p r o t h a l l i a
grown on m ed ia 5 and 6 a r e m ore f i la m e n to u s th a n p l a t e - l i k e tw e n ty - f iv e
d ay s a f t e r g e r m in a t io n . P r o t h a l l i a grow n on th e s e m ed ia h av e
a p p ro x im a te ly t h e same c e l l num ber a s medium 7 , b u t t h e c e l l s a r e
s l i g h t l y s m a l l e r . The d e c re a s e i n am ount o f g ro w th on t h e s e m ed ia from
t h a t on medium 1 was shown t o b e a l a c k o f p h o s p h a te . T h e r e f o r e 5 th e
s m a l le r i n c r e a s e s i n c e l l s i z e and num ber can be a t t r i b u t e d t o t h e l a c k
o f p h o s p h a te . B u t , s in c e t h e p r o t h a l l i a l w e ig h ts f o r b o th f e r n s w ere
g r e a t e r on medium 7 th a n on m ed ia 5 and 6 , i t may b e c o n c lu d e d t h a t s u l f a t e
a l s o can in f lu e n c e c e l l e n la r g e m e n t .
P l a t e 6 o f F ig u re 6 shows W. c o c h i s e n s i s p r o t h a l l i a on medium
10 t h i r t y - f i v e d ay s a f t e r g e r m in a t io n . P r o t h a l l i a from medium 11 w ere
n o t fo u n d t o b e s i g n i f i c a n t l y d i f f e r e n t from th o s e on medium 1 0 .
In t h e a b se n c e o f p h o s p h a te and s u l f a t e t h e p r o t h a l l i a f a i l e d t o d e v e lo p
a p l a t e l e t o r f i l a m e n t . The c e l l num ber i n p r o t h a l l i a o f b o th s p e c ie s
grow n on m ed ia 10 and 11 i s n o t s i g n i f i c a n t l y d i f f e r e n t from t h a t o f
p r o t h a l l i a grow n on medium 7 , b u t th e c e l l s w ere v e ry s m a l l . T h is a g re e s
w i th a c o n c lu s io n d e r iv e d i n t h e p r e v io u s p a r a g r a p h ; s u l f a t e can in f l u e n c e
c e l l e n la r g e m e n t .
I n summ ary, th e p h o to g ra p h ic s t u d i e s and m ic ro s c o p ic o b s e r v a t io n s
h av e b e e n u s e d t o show t h a t when p r o t h a l l i a o f b o th f e r n s a r e grown on
m ed ia c o n ta in in g a d e q u a te p h o s p h a te , t h e c e l l s i z e and num ber d o es n o t
d e c r e a s e . The a d d i t i o n o f s u l f a t e t o a p h o s p h a te - s u l f a t e d e f i c i e n t
medium a l s o c a u se d th e c e l l s i z e t o b e i n c r e a s e d .
![Page 33: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/33.jpg)
36
32
28
24
2 0
16
12
8
4
2. T
24
A————A
A..........A
MEDIUM123456
20 25I
30
DAYS AFTER GERMINATION
Pr«h weight of N . co ch iten tit grown on vorioue omounti of phoephote.
xxitoin 4 .4 mM K , 4 .2 mM C o , 2.1 mM M g, 8 .4 mM N O and 1 .6 mM C l . The N o , HPO , and SO con- ) of the media ore respectively : Medium I , 1 .2 , 1 .4 , 2 .1 j Medium 7 , 1 .0.1 .2 , 2 .4 ; Medium 3 , 0 .^ , 0 .8 , i, 0 .6 , 0 .6 , 2 .9 ; Medium 5 , 0 .4 , 0 .3 , 3 .3 ; Medium 6 , 0 .2 , 0 .0 , 3 .5 .
![Page 34: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/34.jpg)
25
MEDIUM
DAYS AFTER GERMINATION
Figure 3 . The fresh w i e ^ t o f N . sinuota grown on various amounts of phosphate.
All media contain 4 .4 mM K , 4 .2 mM C o , 2.1 mM Mg , 8 . 4 mM N O and 1 .6 mM C l . The N o , HPO , and SO 4 concentrations (mM) of the medio ore respectively: Medium 1, 1 .2 , 1 .4 , 2 .1 ; Medium 2 , 1.0,1 .2 , 2 .4 ; Medium 3 , 0 . 8 , 0 . 8 , 2 .7 ; Medium 4 , 0 . 6 , 0 . 6 , 2 .9 ; Medium 5 , 0 . 4 , 0 .3 , 3 . 3 . ; Medium 6 , 0 . 2 , 0 . 0 , 3 .3 .
![Page 35: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/35.jpg)
26
MEDIUM
DAYS AFTER GERMINATION
Figure 4 . The fresh weight of hi . cochise n»ii grown on modio contoining o def iciency or on excess of phosphate or sulfate .
Medio 7 , 8 , 9 , and 10 contain the following concentrations of ions (mM) respectively: N o , 4.1 , 3 . 5 , 1 .5 , 0 .1 ; K , 8 .0 , 4 . 4 , 7 .2 ,"2 .9^ C o , 4 . 0 , 4 . 2 , 4 . 2 , 4 .2 ; Mg, 4 .2 , 2.1 , 2 . 1 , 3 .4 ; N O , 0 . 2 , 8 . 4 , 8 . 4 , 8 .4 ; C l , 4 . 0 , 1 .6 , 1 .6 , 9 . 7 . The HFO^ and SO concentrations of the media (mM) ore: 0 . 0 , 1 .4 , 1 .4 , 0 .0 and 12 .2 , 3 . 5 , 3 . 5 , 0 . 0 . Medium contains no nutrients. No growth curves were obtained for medio 10 and 11 .
![Page 36: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/36.jpg)
27
MEDIUM
IS 2 0 2 5 SO 3 5
DAYS AFTER GERMINATION
Figure 5 . The fresh weight o f N . t inuoto grown on medio containing a def iciency or an excew of phosphate or su lfa te .
Media 7 , 8 , 9 , and | 0 contain the following concentrations of ions (mM) respectively: N o , 4 . 1 , 3 . 5 , 1 .5 , 0 .1 ; K , 8 . 0 , 4 . 4 , 7 . 2 , 2 .9 ; C a , 4 . 0 , 4 . 2 , 4 . 2 , 4 .2 ; Mg, 4 . 2 , 2 .1 , 2 . 1 , 3 .4 ; N O , 0 . 2 , 8 . 4 , 8 . 4 , 8 .4 ; C l , 4 . 0 , 1 .6 , 1 .6 , 9 . 7 . The H P O j and SO concentrations of the media (mM) are: 0 . 0 , 1 . 4 , 1 0 . 0 , and 1 2 .2 , 3 . 5 , 3 . 5 , 0 . 0 . MediumII contains no nutr ients . Vio growth curves were obtained for media 10 and 11.
![Page 37: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/37.jpg)
Plate 1 .
Plate 2.
Figure 6.
\
28
N . cochisensis prothallium on medium 1 twenty-five days after germination.
oN . cochisensis prothallium on medium 4 twenty-five days after germination.
Plates of N . cochisensis and N . s inuata prothallia grown on different agar media.
I
![Page 38: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/38.jpg)
29
Plate 3. N . sinuata prothallia on medium 4 twenty-five days after germination.
Plate 4 . N . cochisensis prothal lia on medium 7 twenty-five days after germination.
Figure 6 . — Continued.
![Page 39: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/39.jpg)
30
Plate 5.
Plate 6.
N . cochisensis prothallia on medium 5 twenty-five days after germination.
N . cochisensis prothal lia on medium 1 0 thir ty-f ive days after germination .
Figure 6 . — Continued.
![Page 40: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/40.jpg)
SUMMARY
P r o t h a l l i a o f H o th o la e n a c o c h i s e n s i s and H o th o la e n a s i n u a t a
f e r n s w ere grown from s p o re s and c u l t u r e d on a g a r m ed ia t h a t c o n ta in e d
v a r io u s l e v e l s o f p h o s p h a te and s u l f a t e . P h o to g ra p h ic and f r e s h
w e ig h t s t u d i e s w ere u s e d t o m easu re t h e i r g ro w th r e s p o n s e s .
U s in g th e s e s t u d i e s , i t was p o s s i b l e t o show t h a t t h e lo w e s t
l e v e l o f p h o s p h a te r e q u i r e d b y th e f e r n s i s 0 .8 mM i n t h e p r e s e n c e o f
2 .7 mM s u l f a t e and th e o t h e r io n s o f K n o p 's s o l u t i o n .
When p r o t h a l l i a o f b o th f e r n s w ere grown on m ed ia w i th low
am ounts o f p h o s p h a te , t h e c e l l num ber and s i z e w ould d e c r e a s e .
The a d d i t i o n o f s u l f a t e t o p h o s p h a te - s u l f a t e d e f i c i e n t m ed ia w ould
i n c r e a s e th e c e l l s i z e .
I n c o n c lu d in g , H ev ly (5? 6 ) r e p o r t e d t h a t N. c o c h i s e n s i s w ould
grow e q u a l ly w e l l on K n o p 's and P r a n t i ' s m e d ia ; h o w e v e r, N. s i n u a t a
w ould grow w e l l on K n o p 's medium b u t w ould n o t grow on P r a n t l ' s medium.
T hese r e s u l t s were, n o t v e r i f i e d ; i t was fo u n d t h a t W. c o c h i s e n s i s d id
n o t grow e q u a l ly w e l l on b o th m e d ia , and K. s in u a t a w ould grow f o r a t
l e a s t two m onths i f p l a c e d on P r a n t l ' s m edium . The r e s u l t s o b ta in e d i n
t h i s s tu d y do n o t r u l e o u t th e p o s s i b i l i t y t h a t s u l f a t e can s u b s t i t u t e
f o r p h o s p h a te ; i n f a c t , th e y s u g g e s t t h a t s u l f a t e can s u b s t i t u t e f o r
p h o s p h a te , b u t f u r t h e r w ork i s n e e d ed t o e s t a b l i s h t h i s h y p o th e s i s a s
a f a c t .
31
![Page 41: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/41.jpg)
LITERATURE CITED
1 . B a n d u r s k i , R . S . , L, G. W ilso n , and C. L. S q u i r e s . 1 9 5 6 .
The m echanism o f a c t i v e s u l f a t e f o rm a t io n . J . Am er. Chem.
S o c . 7 8 :6 4 0 8 -9 .
2 . Brown, D. 1920 . The. v a lu e o f n u t r i e n t s o lu t i o n s a s c u l t u r e
m ed ia f o r f e r n p r o t h a l l i a . T o rre y a 2 0 :7 6 -8 3 .
3- C o le , Y . , C . L e s a in t , and M. G ra n d je a n . 1963 . The i n f lu e n c e
o f p h o sp h o ru s d e f i c i e n c y on th e m in e r a l c o m p o s itio n o f b a r l e y
a t h a r v e s t . Ann. P h y s io l . V e g e ta b le 5 ( 4 ) :2 7 7 - 8 0 . Chem.
A b s t r . 6 0 : 15089h . 1 $ 6 3 .
4 . F u j iw a r a , A . , and K. T o r i i . 1961 .. P h y s io lo g y o f s u l f a t e on
h ig h e r p l a n t s . I . E f f e c t o f s u l f u r d e f i c i e n c y on m e ta b o lis m
o f h ig h e r p l a n t s . Tohoku J . A gr. R es . 12 ( 3 ) :2 7 7 - 9 0 .
B i o l . A b s t r . 39 ( 1 ):6 1 9 # 7 7 2 0 . 1962 /
5 . . H e v ly , R. H. 1959 . The taxonom y o f t h e N o th o la e n a s i n u a t a
com plex . M a s t e r 's t h e s i s . U n i v e r s i t y o f A r iz o n a , T u cso n .
6. Hevly, R. H. 1963. Adaptations of cheilanthoid ferns to
desert environments. J . Ariz. Acad. Sci. 2 (4):164-75.
7 . K i t s o n , R. E . , and M. G. M e llo n . 1944 . C o lo r im e t r ic d e te r m in a t io n
o f p h o sp h o ru s a s m o ly b d iv a n a d o p h o sp h o r ic a c id . .1 & E Chem.
(A n a l. E D .) 16 (6 ) :3 7 9 - 8 2 .
8 . K u r tz , E . B . , and A. M. Mac Ewan. 1954. I n t r o d u c to r y P l a n t
P h y s io lo g y L a b o ra to ry M anual, 2nd e d . B u r g e s s , M in n e a p o l is .
32
![Page 42: THE PHOSPHATE REQUIREMENT OF THE FERNS byarizona.openrepository.com/arizona/bitstream/10150/... · · 2015-10-22THE PHOSPHATE REQUIREMENT OF THE FERNS NOTHOLAENA COCHISENSIS AND](https://reader031.vdocuments.site/reader031/viewer/2022030416/5aa2541c7f8b9aa0108d118c/html5/thumbnails/42.jpg)
9 . K u r tz , E . B . , and H. B. W aldoiu- i 960 . A new r o l e f o r s u l f u r
In h ig h e r p l a n t s . J . A r iz . A cad . S c i . 1 :1 0 2 -3 .
1 0 . ham m erink , J . 1962 . I n t e r a c t i o n s o f p h o s p h a te and s u l f a t e
on i r r i g a t e d p a s t u r e . New Z e a la n d J . A gr. R e s . 5 :k 9 0 -9 .
1 1 . S te e v e s , T . A ., I . M. S u s s e x , and C . R. P a r ta n e n , 1955 .
In v i t r o s tu d i e s on a b n o rm a l g ro w th o f p r o t h a l l i a o f th e
b r a c k e n f e r n . Am. J . B o t. 4 2 :2 3 2 -4 4 .
1 2 . T a k a h a s h i , N . , T . Yamada, T . H a sh im o to , and J . Yamaki. 1962 .
E f f e c t o f i n o r g a n ic compounds on d a rk g e rm in a t io n o f to b a c c o
s e e d s in d u c e d b y g i b b e r e l l i n . B o t . Mag. 7 5 :4 9 -5 5 . - I n t i . A b s t r .
B i o l . S c i . 2 8 :5 9 6 . 19 6 3 .
1 3 . W ald , G. 1963 . - I n : H o r iz o n s i n B io c h e m is t r y . M. K asha and
B-.' P u llm a n , e d s . p . 1 2 7 -4 2 . Academ ic P r e s s , H.Y. ••
1 4 . ' W h i t t i e r , D. P . I g 6 4 , The e f f e c t o f s u c ro s e on apogamy i n
C yrotom ium f a lc a tu m p r e s l . Am. F e rn J , 5 4 :2 0 -2 4 .